Academic Journal
High taurocholic acid concentration induces ferroptosis by downregulating FTH1 expression in intrahepatic cholestasis of pregnancy
| Title: | High taurocholic acid concentration induces ferroptosis by downregulating FTH1 expression in intrahepatic cholestasis of pregnancy |
|---|---|
| Authors: | Wei-jian Zeng, Hua-jing Yang, Ying-jie Gu, Meng-nan Yang, Meng-ru Sun, Sheng-kai Cheng, Yan-yan Hou, Wei Gu |
| Source: | BMC Pregnancy and Childbirth, Vol 25, Iss 1, Pp 1-13 (2025) |
| Publisher Information: | BMC, 2025. |
| Publication Year: | 2025 |
| Collection: | LCC:Gynecology and obstetrics |
| Subject Terms: | Ferroptosis, FTH1, Intrahepatic cholestasis of pregnancy, Placenta, Taurocholic acid, Gynecology and obstetrics, RG1-991 |
| More Details: | Abstract Background Intrahepatic cholestasis of pregnancy (ICP) is the most common liver disorder associated with pregnancy and is usually diagnosed based on high serum bile acid. However, the pathogenesis of ICP is unclear. Ferroptosis has been reported as an iron-dependent mechanism of cell death. Although the role of Ferritin Heavy Chain 1 (FTH1) in ferroptosis has been extensively studied in various diseases, its mechanism in ICP through ferroptosis is yet to be analyzed. Methods Placental tissues from patients with ICP and healthy controls were employed to verify the expression of FTH1. Taurocholic acid (TCA)-induced HTR-8/SVneo cells were established as an in vitro model for ICP, and ferroptosis-related experiments were performed. In particular, HTR-8/SVneo cells with or without overexpressing FTH1 and HTR-8/SVneo cells with or without TCA induction were investigated to explore the relationship between FTH1 and ferroptosis during ICP in vitro, respectively. Results FTH1 was significantly downregulated in the ICP group compared with the control group. Furthermore, FTH1 and ferroptosis-related protein levels were downregulated, while the intracellular iron, reactive oxygen species, and lipid peroxidation levels were upregulated in the TCA-induced HTR-8/SVneo cells. In contrast, ferroptosis was inhibited by overexpression of FTH1 in TCA-induced HTR-8/SVneo cells. Conclusions A high concentration of TCA in HTR-8/SVneo cells decreased the expression of FTH1. Overexpression of FTH1 could prevent cell death from ferroptosis induced by TCA. Thus, inhibiting the downregulation of FTH1 could be a potential therapeutic target for ICP treatment. |
| Document Type: | article |
| File Description: | electronic resource |
| Language: | English |
| ISSN: | 1471-2393 |
| Relation: | https://doaj.org/toc/1471-2393 |
| DOI: | 10.1186/s12884-025-07143-9 |
| Access URL: | https://doaj.org/article/f93066ee320141cf9cb53e89f1e12d49 |
| Accession Number: | edsdoj.f93066ee320141cf9cb53e89f1e12d49 |
| Database: | Directory of Open Access Journals |
| Full text is not displayed to guests. | Login for full access. |
| FullText | Links: – Type: pdflink Url: https://content.ebscohost.com/cds/retrieve?content=AQICAHjPtM4BHU3ZchRwgzYmadcigk49r9CVlbU7V5F6lgH7WwG19ZSU08vvo9Nepo77gzKSAAAA4TCB3gYJKoZIhvcNAQcGoIHQMIHNAgEAMIHHBgkqhkiG9w0BBwEwHgYJYIZIAWUDBAEuMBEEDCu1XeDh5rsAJ8pvlgIBEICBmXlvil3oTn49tu-K-QS_yDaORb3--bOboBhmQch_6WJhpy1WD0XwJ9HbNMA1MPGjfeqwWvNlxOOICoJz8srQfmdp81nJmLBJR_7ffGn5jb9wUqR6F2cuJuHwdEvcgSqvJtBlNiM8tt2Lo30Fin5L_r_khQ34BvVZgIPDoJsPDbADsqYmYJYB6P56ykdjd-OGnm-jOUAmXcpNZg== Text: Availability: 1 Value: <anid>AN0182154229;[1cii]09jan.25;2025Jan13.02:39;v2.2.500</anid> <title id="AN0182154229-1">High taurocholic acid concentration induces ferroptosis by downregulating FTH1 expression in intrahepatic cholestasis of pregnancy </title> <p>Background: Intrahepatic cholestasis of pregnancy (ICP) is the most common liver disorder associated with pregnancy and is usually diagnosed based on high serum bile acid. However, the pathogenesis of ICP is unclear. Ferroptosis has been reported as an iron-dependent mechanism of cell death. Although the role of Ferritin Heavy Chain 1 (FTH1) in ferroptosis has been extensively studied in various diseases, its mechanism in ICP through ferroptosis is yet to be analyzed. Methods: Placental tissues from patients with ICP and healthy controls were employed to verify the expression of FTH1. Taurocholic acid (TCA)-induced HTR-8/SVneo cells were established as an in vitro model for ICP, and ferroptosis-related experiments were performed. In particular, HTR-8/SVneo cells with or without overexpressing FTH1 and HTR-8/SVneo cells with or without TCA induction were investigated to explore the relationship between FTH1 and ferroptosis during ICP in vitro, respectively. Results: FTH1 was significantly downregulated in the ICP group compared with the control group. Furthermore, FTH1 and ferroptosis-related protein levels were downregulated, while the intracellular iron, reactive oxygen species, and lipid peroxidation levels were upregulated in the TCA-induced HTR-8/SVneo cells. In contrast, ferroptosis was inhibited by overexpression of FTH1 in TCA-induced HTR-8/SVneo cells. Conclusions: A high concentration of TCA in HTR-8/SVneo cells decreased the expression of FTH1. Overexpression of FTH1 could prevent cell death from ferroptosis induced by TCA. Thus, inhibiting the downregulation of FTH1 could be a potential therapeutic target for ICP treatment.</p> <p>Keywords: Ferroptosis; FTH1; Intrahepatic cholestasis of pregnancy; Placenta; Taurocholic acid</p> <p>Supplementary Information The online version contains supplementary material available at https://doi.org/10.1186/s12884-025-07143-9.</p> <hd id="AN0182154229-2">Background</hd> <p>Intrahepatic cholestasis of pregnancy (ICP), also known as gestational cholestasis, is defined as the presence of pruritus and elevated levels of serum total bile acid (TBA) and liver transaminases [[<reflink idref="bib1" id="ref1">1</reflink>], [<reflink idref="bib3" id="ref2">3</reflink>], [<reflink idref="bib5" id="ref3">5</reflink>]–[<reflink idref="bib6" id="ref4">6</reflink>]]. ICP is the most common liver disease associated with pregnancy [[<reflink idref="bib2" id="ref5">2</reflink>], [<reflink idref="bib4" id="ref6">4</reflink>], [<reflink idref="bib5" id="ref7">5</reflink>]–[<reflink idref="bib6" id="ref8">6</reflink>]]. Notably, the incidence of ICP has obvious regional and ethnic differences, with the lowest rate found in Europe (0.1‒0.2%) and the highest rate found in South America (9.2‒15.6%) [[<reflink idref="bib6" id="ref9">6</reflink>]]. Chile, Sweden, and the Yangtze River Basin in China also have a higher incidence of ICP relative to other countries. The etiology of ICP is unclear but considered to be related to female hormones, genetics, immune, and environmental factors [[<reflink idref="bib3" id="ref10">3</reflink>], [<reflink idref="bib4" id="ref11">4</reflink>]–[<reflink idref="bib5" id="ref12">5</reflink>], [<reflink idref="bib7" id="ref13">7</reflink>]]. Although the maternal symptoms of ICP may spontaneously disappear soon after postpartum, ICP can cause serious consequences, such as sudden intrauterine fetal demise, fetal distress, preterm birth, and slower growth of offspring [[<reflink idref="bib5" id="ref14">5</reflink>], [<reflink idref="bib8" id="ref15">8</reflink>]]. In recent years, a high concentration of TBA was considered to be a crucial factor in trophoblast dysfunction and the etiology of fetal complications in patients with ICP [[<reflink idref="bib10" id="ref16">10</reflink>]]. Taurocholic acid (TCA) is one of the main bile acids whose levels are elevated in these patients and may be involved in the pathogenesis of ICP [[<reflink idref="bib11" id="ref17">11</reflink>]]. Thus, TCA-induced human trophoblast cells have been widely used as an in vitro model for the investigation of ICP.</p> <p>Ferroptosis represents a distinct type of iron-dependent cell demise, exhibiting notable differences in morphology, biochemistry, and genetics when compared to apoptosis, necrosis, and autophagy [[<reflink idref="bib12" id="ref18">12</reflink>]]. It has been reported as a prominent area of investigation in various diseases, such as brain injury, heart injury, breast cancer, asthma, and preeclampsia [[<reflink idref="bib13" id="ref19">13</reflink>], [<reflink idref="bib14" id="ref20">14</reflink>]–[<reflink idref="bib15" id="ref21">15</reflink>]]. Ferritin Heavy Chain 1 (FTH1), plays a key role in ferroptosis, a key subunit of the ferritin 'nanocage' that stores iron in its non-toxic ferric form, plays an important role in the maintenance of iron homeostasis in cells [[<reflink idref="bib16" id="ref22">16</reflink>], [<reflink idref="bib17" id="ref23">17</reflink>]–[<reflink idref="bib18" id="ref24">18</reflink>]]. Thus, FTH1 can prevent the harmful effects induced by iron overload. The function of FTH1 in the pathogenesis of various diseases, such as cancer [[<reflink idref="bib19" id="ref25">19</reflink>]], myocardial injury and cardiac dysfunction [[<reflink idref="bib21" id="ref26">21</reflink>]], and cerebral ischemia-reperfusion injury [[<reflink idref="bib22" id="ref27">22</reflink>]], was recently reported. Our previous study of proteomics analysis revealed the downregulation of FTH1 in the placental tissues in the ICP group compared with control groups [[<reflink idref="bib23" id="ref28">23</reflink>]]. The exact mechanism and role of FTH1 on ferroptosis in the placenta of patients with ICP have not been reported. We hypothesized that the downregulation of FTH1 in patients with ICP could be a key point for elucidating the mechanism involved in ferroptosis.</p> <hd id="AN0182154229-3">Methods</hd> <p></p> <hd id="AN0182154229-4">Patients and sample collection</hd> <p>A total of 20 patients who underwent cesarean section at the obstetric department of the International Peace Maternity and Child Health Hospital (IPMCH) between March 16, 2020, and September 17, 2020, were invited to participate in this study. Patients with singleton pregnancy who had regular antenatal examinations were divided into the ICP (<emph>n</emph> = 10) and control groups (<emph>n</emph> = 10). ICP is defined as the presence of pruritus and elevated levels of serum TBA and liver transaminases. The inclusion criteria for this study were (<reflink idref="bib1" id="ref29">1</reflink>) Chinese nationality, (<reflink idref="bib2" id="ref30">2</reflink>) singleton pregnancy, (<reflink idref="bib3" id="ref31">3</reflink>) diagnosed with ICP, (<reflink idref="bib4" id="ref32">4</reflink>) delivered at IPMCH, and (<reflink idref="bib5" id="ref33">5</reflink>) signed informed consent forms. Participants were excluded if they were diagnosed with any one of the diseases including gestational diabetes mellitus, chronic hypertension, renal disease, thyroid dysfunction, and pregnancy with malignant tumor.</p> <p>The detailed patient characteristics are outlined in Table 1. Placental tissue samples were collected from all enrolled individuals. These samples were extensively rinsed with PBS and sterile water and divided into two portions: one portion was snap-frozen samples in liquid nitrogen and stored at − 80 ℃ for further testing, and the other portion was fixed with formalin and then embedded in paraffin for immunohistochemical staining analysis.</p> <p>Table 1 Clinical characteristics of the included subjects</p> <p> <ephtml> &lt;table frame="hsides" rules="groups"&gt;&lt;thead&gt;&lt;tr&gt;&lt;th align="left" /&gt;&lt;th align="left"&gt;&lt;p&gt;Control(&lt;italic&gt;n&lt;/italic&gt; = 10)&lt;/p&gt;&lt;/th&gt;&lt;th align="left"&gt;&lt;p&gt;ICP(&lt;italic&gt;n&lt;/italic&gt; = 10)&lt;/p&gt;&lt;/th&gt;&lt;th align="left"&gt;&lt;p&gt;&lt;italic&gt;p&lt;/italic&gt; value&lt;/p&gt;&lt;/th&gt;&lt;/tr&gt;&lt;/thead&gt;&lt;tbody&gt;&lt;tr&gt;&lt;td align="left"&gt;&lt;p&gt;Maternal age (y)&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;32.1 &amp;#177; 5.2&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;31.9 &amp;#177; 2.9&lt;/p&gt;&lt;/td&gt;&lt;td char="." align="char"&gt;&lt;p&gt;0.917&lt;/p&gt;&lt;/td&gt;&lt;/tr&gt;&lt;tr&gt;&lt;td align="left"&gt;&lt;p&gt;Maternal BMI (kg/m&amp;#178;)&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;20.9 &amp;#177; 0.9&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;19.5 &amp;#177; 1.3&lt;/p&gt;&lt;/td&gt;&lt;td char="." align="char"&gt;&lt;p&gt;0.013&lt;/p&gt;&lt;/td&gt;&lt;/tr&gt;&lt;tr&gt;&lt;td align="left"&gt;&lt;p&gt;Gestational age at delivery(week)&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;38.8 &amp;#177; 0.4&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;37.3 &amp;#177; 1.2&lt;/p&gt;&lt;/td&gt;&lt;td char="." align="char"&gt;&lt;p&gt;0.001&lt;/p&gt;&lt;/td&gt;&lt;/tr&gt;&lt;tr&gt;&lt;td align="left"&gt;&lt;p&gt;Newborn birth weight (g)&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;3223.5 &amp;#177; 296.4&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;3044.5 &amp;#177; 233.8&lt;/p&gt;&lt;/td&gt;&lt;td char="." align="char"&gt;&lt;p&gt;0.151&lt;/p&gt;&lt;/td&gt;&lt;/tr&gt;&lt;tr&gt;&lt;td align="left"&gt;&lt;p&gt;TBA(&amp;#181;mol/L)&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;3.3 &amp;#177; 1.4&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;37.9 &amp;#177; 8.3&lt;/p&gt;&lt;/td&gt;&lt;td char="." align="char"&gt;&lt;p&gt;&amp;#60;0.001&lt;/p&gt;&lt;/td&gt;&lt;/tr&gt;&lt;tr&gt;&lt;td align="left"&gt;&lt;p&gt;TBIL(&amp;#181;mol/L)&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;4.8 &amp;#177; 1.9&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;5.1 &amp;#177; 0.9&lt;/p&gt;&lt;/td&gt;&lt;td char="." align="char"&gt;&lt;p&gt;0.665&lt;/p&gt;&lt;/td&gt;&lt;/tr&gt;&lt;tr&gt;&lt;td align="left"&gt;&lt;p&gt;DBIL(&amp;#181;mol/L)&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;2.4 &amp;#177; 0.6&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;2.4 &amp;#177; 0.6&lt;/p&gt;&lt;/td&gt;&lt;td char="." align="char"&gt;&lt;p&gt;0.798&lt;/p&gt;&lt;/td&gt;&lt;/tr&gt;&lt;tr&gt;&lt;td align="left"&gt;&lt;p&gt;ALT (U/L)&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;9.1 &amp;#177; 1.5&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;7.3 &amp;#177; 2.4&lt;/p&gt;&lt;/td&gt;&lt;td char="." align="char"&gt;&lt;p&gt;0.058&lt;/p&gt;&lt;/td&gt;&lt;/tr&gt;&lt;tr&gt;&lt;td align="left"&gt;&lt;p&gt;AST (U/L)&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;13.7 &amp;#177; 1.6&lt;/p&gt;&lt;/td&gt;&lt;td char="&amp;#177;" align="char"&gt;&lt;p&gt;13.7 &amp;#177; 2.4&lt;/p&gt;&lt;/td&gt;&lt;td char="." align="char"&gt;&lt;p&gt;1.000&lt;/p&gt;&lt;/td&gt;&lt;/tr&gt;&lt;/tbody&gt;&lt;/table&gt; </ephtml> </p> <p>ICP: intrahepatic cholestasis of pregnancy, BMI: body mass index, TBA: total bile acid, TBIL: total bilirubin, DBIL: direct bilirubin, ALT: serum alanine transaminase, AST: serum aspartate transaminase</p> <hd id="AN0182154229-5">RT-qPCR and western blot</hd> <p>RT-qPCR and western blot analysis were performed as previously described [[<reflink idref="bib24" id="ref34">24</reflink>]]. Total RNA of placental tissues (Control groups, n = 8; ICP groups, n = 8) was extracted with TRIzol reagent (Invitrogen) and was dissolved in 10 µL of RNase-free water. Complementary DNA (cDNA) was synthesized utilizing the EntiLink™ 1st Strand cDNA Synthesis Kit (ELK Biotechnology), and real-time qPCR was conducted to assess gene expression utilizing EnTurbo™ SYBR Green PCR SuperMix (ELK Biotechnology) on a StepOne™ Real-time PCR system (Life Technologies). The expression of genes was normalized to that of GAPDH. The primer sequences for GAPDH were sense: 5'-CATCATCCCTGCCTCTACTGG-3' and antisense: 5'-GTGGGTGTCGCTGTTGAAGTC-3'. The primer sequences for FTH1 were sense 5'-CCCCCATTTGTGTGACTTCAT-3' and antisense 5'-GCCCGAGGCTTAGCTTTCATT-3'. The results are representative of three independent experiments.</p> <p>Sixteen placental tissues (Control groups, <emph>n</emph> = 8; ICP groups, <emph>n</emph> = 8) were lysed with RIPA lysis buffer (ASPEN) in the presence of protease inhibitor, and then the isolated protein was quantified and evaluated utilizing using the BCA Protein Assay Kit (Pierce, Rockford, IL, USA). 15 µg of protein lysate was loaded onto sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to polyvinylidene difluoride membranes. Non-specific binding was blocked with 5% skimmed milk for 1 h at 20 ℃. Next, the membranes were incubated with anti-FTH1 (1:1000; Abcam, Cat #: ab65080) as the primary antibody at 4℃ overnight, respectively, followed by a secondary antibody (1:5000, Affinity, Shanghai, China) at 20 ℃ for 30 min. anti-α-tubulin primary antibody (1:10,000; Abcam, Cat #:ab52866) was used as an internal control. The antigens and antibodies were detected using high-signal ECL substrates, and the gray values of protein bands were analyzed and quantified using Image J (NIH, version 1.47). For the cell samples the following primary antibodies: anti-SLC7A11 (1: 3000; Abcam, Cat #: ab175186), anti-GPX4 (1:1000; Abcam, Cat #: ab125066), anti-FTH1 (1:1000; Abcam, Cat #: ab65080), anti-GAPDH (1:1000; CST, Cat #: 2118), anti-KRAS(1:1000; Abcam, Cat #:ab275876) were used.</p> <p> <bold>Immunohistochemical (IHC) staining</bold>.</p> <p>Immunohistochemical staining was carried out with twenty placental tissues (Control groups, <emph>n</emph> = 10; ICP groups, <emph>n</emph> = 10) as described previously [[<reflink idref="bib24" id="ref35">24</reflink>]]. Paraffin-embedded placental tissue sections (thickness = 3 μm) were deparaffinized and rehydrated in aqueous ethanol solutions of different concentrations, followed by treatment with citric acid buffer to extract antigens. After washing with phosphate-buffered saline (PBS), endogenous peroxidase was blocked by incubating with 3% H<subs>2</subs>O<subs>2</subs> for 10 min. PBS containing 10% goat serum was added to block non-specific binding. Next, the sections were incubated with anti-FTH1 (1:1000, Abcam, ab92478) overnight at 4 ℃, followed by a secondary antibody for 30 min at 20 ℃. The samples were incubated with an avidin-biotin complex reagent to detect the bound antibody, followed by diaminobenzidine solution for 2 h. The sections were then re-stained with hematoxylin. After rinsing and air-drying, the samples were sealed with neutral resin. The full staining slides were scanned at the NIEHS Image Core. Image-Pro plus 6.0 software (Media Cybernetics, Inc. Marlow, UK) was used to assess the area and density of the dyed region, and the integrated optical density (IOD) value of the IHC section. The mean densitometry of the digital image (magnification, ×400) was designated as representative FTH1 staining intensity. The signal density of the tissue areas from five randomly selected fields was counted in a blinded manner and subjected to statistical.</p> <hd id="AN0182154229-6">Bioinformatics analyses</hd> <p>The differentially expressed proteins in our previous study were employed for upstream regulator analysis via Qiagen's Ingenuity Pathway Analysis (IPA, version 01–13, QIAGEN Redwood City, CA, <ulink href="http://www.Ingenuity.com">www.Ingenuity.com</ulink>) with default setting [[<reflink idref="bib23" id="ref36">23</reflink>]]. The Regulator Effects network was generated by IPA.</p> <hd id="AN0182154229-7">Cell culture and treatment with taurocholic acid</hd> <p>The human extravillous trophoblast cell line, HTR-8/SVneo, and human immortalized umbilical endothelial cells (EAhy926) were purchased from Shanghai Biowing Applied Biotechnology Co. LTD (Shanghai, China). Additionally, the authentication of both cells was checked by STR profiling (Supplementary files 1 and 2). HTR-8/SVneo cells were maintained in RPMI 1640 medium (Gibco, Cat#:11875093) supplemented with 10% heat-inactivated FBS and 1% penicillin-streptomycin at 37 ℃ in a 5% CO<subs>2</subs> incubator. EAhy926 cells were maintained in DMEM/F12 medium (Gibco, Cat#: C11330500BT) supplemented with 10% heat-inactivated FBS at 37 ℃ in a 5% CO<subs>2</subs> incubator. Cells were seeded in 6-well plates and grown until 80% confluency. Thereafter, 10, 50, and 100 µM TCA (Sigma-Aldrich, Cat #: T4009) were added to the culture medium to represent the serum bile acid levels observed in normal pregnancy, and moderate and severe cholestasis, respectively [[<reflink idref="bib25" id="ref37">25</reflink>]]. The cells treated with an equal volume of vehicle (distilled water) in the medium were used as a negative control (0 µM).</p> <hd id="AN0182154229-8">MTT assay</hd> <p>The assay was performed using MTT Based Cell Growth Determination Kit (Sigma-Aldrich, Cat #: CGD-1) with slight modifications [[<reflink idref="bib26" id="ref38">26</reflink>]]. Briefly, a total of 5 × 10<sups>3</sups> cells were seeded in each well on 96-well plates overnight. Then the cells treated with different concentrations of TCA (0, 10, 50, and 100 µM) for different exposure times (<reflink idref="bib6" id="ref39">6</reflink>, 12, 24, and 48 h) were collected for assay. 10 µL MTT (methylthiazol tetrazolium) solution was added into each well for incubation at 37 ℃ for 4 h. The absorbance (optical density; OD) values at 570 nm were measured using a microplate reader (Infinite 200 PRO, Tecan Trading AG, Switzerland). The assays were repeated five times.</p> <hd id="AN0182154229-9">Cell viability assay</hd> <p>Cell survival experiments were carried out on 96-well plates using Cell Counting Kit-8 (CCK8) survival assay (Sigma–Aldrich, Cat #: 96992) according to the manufacturer's instructions. Cells were seeded in 96-well plates at a density of 5 × 10<sups>3</sups> cells/mL. Then, the medium was replaced with different treatment groups. After different treatment hours(0, 24, 48, and 72 h), 10 µL of CCK-8 reagent was added to each well and incubated for 1 h. The OD values at 450 nm were recorded on a microplate reader (Infinite 200 PRO, Tecan Trading AG, Switzerland). Each experiment was performed in triplicate.</p> <hd id="AN0182154229-10">Intracellular iron level assay</hd> <p>Intracellular iron levels were determined using an Intracellular Iron Colorimetric Assay Kit (Applygen, Cat#: E1042) according to the manufacturer's recommendations. In brief, 2 × 10<sups>5</sups> cells cultured in 6-well plates were washed with PBS and lysed in 200 µL of lysis buffer containing 1% Triton X-100. The iron standards of different concentrations and working reagents were prepared with the instructions of this Kit. 200 µL of the obtained supernatant from each sample was transferred into a 96-well plate to test the OD values at 550 nm by a microplate reader (Infinite 200 PRO, Tecan Trading AG, Switzerland). Each experiment was performed in triplicate. Results were expressed as means ± SD.</p> <hd id="AN0182154229-11">Intracellular malondialdehyde (MDA) assay</hd> <p>MDA was measured using an MDA Kit (Beyotime Biotech, Cat #: S0131S) according to the manufacturer's recommendations. Briefly, 10<sups>6</sups> cells were collected after different treatments and lysed in 100 µL of lysis buffer containing 1% Triton X-100. Then the cell lysate was centrifuged at 12,000 × g for 10 min. MDA in the sample reacts with thiobarbituric acid (TBA) to generate an MDA-TBA adduct that could be detected at 532 nm by a microplate reader (Infinite 200 PRO, Tecan Trading AG, Switzerland). The protein level was also tested by the BCA Protein Assay Kit (Pierce, Rockford, IL, USA). Results were normalized to protein content in cell lysates and expressed as means ± SD. Each experiment was repeated five times.</p> <hd id="AN0182154229-12">Lipid peroxidation (4-HNE) assay</hd> <p>In brief, 2 × 10<sups>5</sups> cells from different groups were collected for 4-HNE Assay with the 4-HNE Assay Kit (Abcam, Cat #: ab238538) according to the manufacturer's instructions. 50 µL standard or sample were added to the wells of 4-HNE Conjugate coated plate and incubated for 10 min. 50 µL of the diluted anti-4-HNE antibody was added and incubated for 1 h. Followed by the washing step with 250 µL Wash Buffer. Then 100 µL diluted Secondary Antibody-HRP Conjugate was added per well and incubated for another 1 h followed by a washing step. Then 100 µL of warm Substrate Solution was added and incubated for 10 min. Finally, the OD values at 450 nm were tested by a microplate reader (Infinite 200 PRO, Tecan Trading AG, Switzerland). Each experiment was repeated five times. Results were shown as means ± SD.</p> <hd id="AN0182154229-13">Intracellular reactive oxygen species (ROS) assay</hd> <p>The cellular ROS (cROS) in HTR-8/SVneo cells were stained with 2′,7′-dichlorofluorescein diacetate (DCFH-DA, Beyotime Biotechnology, Cat #: S0033). Cells were seeded onto 6-well plates at a density of 2 × 10<sups>5</sups> cells per well and cultured for 24 h. After washing twice with PBS, different groups of treatment were added and cells were incubated for 24 h. Then, 20 µM DCFH-DA was added and cells were incubated for 30 min at 37 ℃. The cells were washed twice with PBS, 400 µL of PBS was added to each well, and fluorescence intensity was determined by flow cytometry (UTHSCSA, Flow Cytometry Core) according to the manufacturer's instructions.</p> <hd id="AN0182154229-14">Mitotracker red CMX ROS staining</hd> <p>To determine the mitochondrial ROS (mROS) levels, HTR-8/SVneo cells were seeded at 2 × 10<sups>5</sups> cells/well in 8-well ibidi plates and subjected to different conditions. Thereafter, different groups of cells were stained with 200 nmol/L MitoTracker Red CMXRos (Thermo Fisher Scientific, Cat#: M36008) for 15 min at 37 °C and then incubated with Hoechst nuclear stain (Dojindo) for 10 min in the dark. The cells were then washed with PBS and randomly captured using a Leica SP8 confocal laser scanning microscope (Leica Microsystems) with a Plan Apochromat 60 × 1.40 NA oil immersion objective. Slices were collected every 1.5 μm to generate z-stacks, and each image represented the maximum projection of all slices along the z-stack. Images were determined across nine randomly selected fields from three independent experiments.</p> <hd id="AN0182154229-15">Cell transfection</hd> <p>HTR-8/SVneo cells were sub-cultured in 6-well plates at a density of 2 × 10<sups>5</sups> cells/well with complete culture medium overnight. The FTH1-overexpressed plasmid (FTH1 overexpression, OE) and its corresponding empty vector plasmids (negative control, NC) were transfected into HTR-8/SVneo cells using lipofectamine 2000 (Invitrogen) according to the manufacturer instructions. After 24 h, the cells were collected to determine the overexpression efficiency. The medium was then exchanged, and the cells were treated with TCA (100 µM) for an additional 24 h. Finally, cells were harvested for western blot and the remaining analyses as described above. The sequences of the FTH1-overexpressed vectors are outlined in supplementary file 3. Herein, 10 µM of deferoxamine (DFO, Abcam, Cat #: ab120727), the ferroptosis inhibitor, was utilized for 24 h. TCA-induced cell co-treatment with DFO was also performed with overexpressing and negative control cells.</p> <hd id="AN0182154229-16">Statistical analysis</hd> <p>Statistical analysis was carried out using Prism 9 from GraphPad Software (San Diego, CA). The data are expressed as means ± SD and compared using the independent samples t-test. One-way ANOVA followed by Fisher's LSD was used for multiple comparisons. Multiple comparisons were performed in cell viability, intracellular iron level, MDA level, and 4-HNE assay. A value of <emph>p</emph> &lt; 0.05 was considered statistically significant.</p> <hd id="AN0182154229-17">Results</hd> <p></p> <hd id="AN0182154229-18">Expression of FTH1 in placenta</hd> <p>Eight placental tissues from the ICP and control groups were randomly selected to verify the expression of FTH1 screened from our previous proteomics study using western blot analysis and RT-qPCR. The western blot results and its relative gray intensity to α-tubulin were lower in the ICP group, including the expression of the FTH1 mRNA compared to that in the control group (Fig. 1A‒C). All placental tissues from the ICP (<emph>n</emph> = 10) and control groups (<emph>n</emph> = 10) were subjected to IHC staining analysis. FTH1 positive cells had brown yellow cytoplasm or nuclear membrane, while FTH1 negative cells had blue-stained nuclei (Fig. 1D). The gray-level histogram of the two groups revealed that the expression of FTH1 was lower in the ICP group compared to that in the control group (Fig. 1E), aligning with our previous result.</p> <p>Graph: Fig. 1 Expression of Ferritin Heavy Chain 1 (FTH1) in the placental tissue of the intrahepatic cholestasis of pregnancy (ICP) and control groups A‒B. Representative protein bands and quantitative analysis of FTH1 protein expression by western blotting. The data represent the mean ± SD. *p &lt; 0.05 vs. Control C. The related expression of mRNA. The data represent the mean ± SD.**p &lt; 0.01 vs. Control. D. Immunohistochemical staining images showing the expression and localization of FTH1 in placental tissues. Scale bar, 100 μm. E. Quantitative analysis of FTH1 expression in the two groups. Quantification of FTH1 staining intensity showed that FTH1expression was significantly decreased in placental tissues of ICP groups compared to those from control groups. The data represent the mean ± SD. **p &lt; 0.01 vs. Control</p> <hd id="AN0182154229-19">TCA induced expression level of FTH1 in cell-line models</hd> <p>To establish an in vitro cellular model of ICP, different concentrations of TCA and different exposure times for HTR-8/SVneo cells and EAhy926 cells were tested using the MTT assay. The OD values represent the amount of surviving cells. Overall, the absorbance values of each group for both cell lines decreased with increased TCA concentrations at the same exposure time. As shown in Fig. 2A, the cell viability decreased with increasing concentrations of TCA in EAhy926 cells. The HTR-8/SVneo cells showed a similar trend, with one exception (Fig. 2B). The cell viability of HTR-8/SVneo cells treated with 100 µM TCA for 48 h increased compared with that of cells treated with 50 µM TCA for 48 h. Overall, TCA-induced cell death of HTR-8/SVneo and EAhy926 cells occurred in a dose-dependent manner. Thus, HTR-8/SVneo cells treated with TCA (100 µM) for 24 h were selected to represent the pathological environment of ICP and used for subsequent studies. The expression of FTH1 was evaluated in the TCA treatment group (100 µM for 24 h) and control group by RT-qPCR and western blot. Interestingly, the protein and mRNA expression of FTH1 were significantly downregulated in the TCA treatment group (Fig. 2C).</p> <p>Graph: Fig. 2 Effect of taurocholic acid (TCA) on the cell viability and expression of FTH1A. Effect of TCA on EAhy926 cells based on the MTT assay. The data represent the mean ± SD, *p &lt; 0.05 vs. Control, **p &lt; 0.01 vs. Control.B. Effect of TCA on the viability of HTR-8/SVneo cells based on the MTT assay. The data represent the mean ± SD, **p &lt; 0.01 vs. Control. C. The protein and mRNA expression levels of FTH1 were downregulated in HTR-8/SVneo cells treated with TCA (100 µM for 24 h). The data represent the mean ± SD, **p &lt; 0.01 vs. Control</p> <hd id="AN0182154229-20">Regulator effects network analysis related to FTH1 by IPA</hd> <p>A total 8 of Regulator Effects networks were enriched by IPA, and the top one was shown in Fig. 3 with the highest Consistency Score 5.965 (Supplementary file 4). The downreguled of FTH1 and the other differentially expressed proteins found in our previous study could be used to predict the activation of organismal death by IPA. GTPase KRas (KRAS) could not only activate organismal death directly, but also activate organismal death by inhibiting the expression of FTH1 through a transcriptional mechanism as shown by IPA.</p> <p>Graph: Fig. 3 Regulator effects network of differentially expressed proteins in the placental tissue of the ICP and control groups via proteomics analysis by IPA. (red = upregulated, blue = downregulated, orange line = predicted activation, blue line = predicted inhibition)</p> <hd id="AN0182154229-21">TCA-induced HTR-8/SVneo cell ferroptosis by downregulating the expression of FTH1</hd> <p>Western blot analysis revealed that the levels of FTH1, glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11(SLC7A11) and KRAS were decreased in TCA-treated HTR-8/SVneo cells (Fig. 4A). To verify that ferroptosis can cause cell death induced by TCA, DFO, a ferroptosis inhibitor, was used. As shown in Fig. 4A, DFO could increase the expression of FTH1, GPX4, and SLC7A11 compared with that found in the control group. DFO could remarkably inhibit ferroptosis, as confirmed by western blot results. FTH1, GPX4, and SLC7A11 were upregulated in the DFO group. TCA-induced cell co-treatment with DFO remarkably rescued TCA-induced cell death (Fig. 4A); however, the expression of KRAS remained downregulated. As shown in Fig. 4B, cell growth was remarkably inhibited in the TCA group compared with that in the control group. The proliferation and viability of cells in the DFO group were only affected at 24 h, and were consistent with those in the control group thereafter. For TCA-induced cell co-treatment with DFO (TCA + DFO group), DFO treatment had partially offset the inhibition of TCA on cells, particularly after 24 h of treatment. The intracellular iron levels were significantly increased in the TCA group (32.29 ± 2.74 µM) compared with those in the control group (17.19 ± 1.28 µM). Despite the co-treatment with DFO, the iron levels markedly decreased (20.79 ± 1.09 µM) (Fig. 4C). Similarly, the MDA levels increased in the TCA group (13.92 ± 0.32 nmol/mg) compared with those in the control group (4.85 ± 0.15 nmol/mg) (Fig. 4D). The MDA level in the TCA + DFO group was 9.01 ± 0.16 nmol/mg, and the 4-HNE results showed a similar trend (Fig. 4E). Also, cROS and mROS levels were measured using DCFH-DA and MitoSOX, respectively, via confocal laser scanning microscopy. ROS levels were quantified using a flow cytometer. As shown in Fig. 4F, the TCA group had the highest ROS level; this level was found to be significant relative to that of the other groups, including the green fluorescence visualized and captured using a confocal microscope (Fig. 4G). Similarly, the red fluorescence of the TCA group was the most obvious among the four groups. DFO could effectively reduce the production of cROS and mROS induced by TCA in cells. Taken together, these findings suggest that TCA triggered ferroptosis of HTR-8/SVneo cells.</p> <p>Graph: Fig. 4 TCA induces the ferroptosis of HTR-8/SVneo cells by downregulating FTH1. A. Expression and quantitative analysis of FTH1, GPX4, SLC7A11, and KRAS based on western blotting. B. HTR-8/SVneo cells were treated with or without TCA (100 µM) or DFO (10 µM) for 24 h, and then the cell viability was measured. Control group: HTR-8/SVneo cells without any treatment; TCA group: HTR-8/SVneo cells treated with 100 µM TCA for 24 h; DFO group: HTR-8/SVneo cells treated with 10 µM DFO for 24 h; TCA + DFO group: HTR-8/SVneo cells co-treated with 100 µM TCA and 10 µM DFO for 24 h, mean ± SD, n = 5; **p &lt; 0.01 vs. Control, ##p &lt; 0.01 vs. TCA. C. Intracellular iron levels in the control, TCA, DFO, and TCA + DFO groups. mean ± SD, n = 5; *p &lt; 0.05 vs. Control, **p &lt; 0.01 vs. Control, ##p &lt; 0.01 vs. TCA. D. MDA levels in the control, TCA, DFO, and TCA + DFO groups. mean ± SD, n = 5; **p &lt; 0.01 vs. Control, ##p &lt; 0.01 vs. TCA. E.4-HNE levels in the control, TCA, DFO, and TCA + DFO groups. mean ± SD, n = 5; **p &lt; 0.01 vs. Control, ##p &lt; 0.01 vs. TCA.F. HTR-8/SVneo cells were treated with or without TCA (100 µM) or DFO (10 µM) for 24 h, and then ROS levels were detected by flow cytometry. G. Representative images of HTR-8/SVneo cells stained with DCFH-DA and MitoSox-Red. Scale bar, 5 μm FTH1: Ferritin Heavy Chain 1, KRAS: GTPase KRas, GPX4: glutathione peroxidase 4, SLC7A11: solute carrier family 7 member 11, TCA: taurocholic acid, DFO: deferoxamine, OD: optical density, MDA: malondialdehyde, 4-HNE: lipid peroxidation, ROS: reactive oxygen species, DCFH-DA:2′,7′-dichlorofluorescein diacetate</p> <hd id="AN0182154229-22">Effect of FTH1 overexpression on TCA-induced HTR-8/SVneo cell ferroptosis</hd> <p>To determine whether FTH1 plays a key role in ferroptosis induced by TCA, HTR-8/SVneo cells were transfected with FTH1 plasmid with or without TCA. TCA-induced cell co-treatment with DFO was also performed. As shown in Fig. 5A, FTH1 was significantly overexpressed at the mRNA and protein levels.</p> <p>Graph: Fig. 5 FTH1 is a key determinant of TCA-induced ferroptosis of HTR-8/SVneo cells. A. The protein and mRNA expression levels of FTH1 were upregulated in HTR-8/SVneo cells following transfection with the FTH1-overexpression plasmid and negative control. **p &lt; 0.01. B. Expression levels of FTH1, GPX4, SLC7A11, and KRAS based on western blotting. C. Untransfected cells and FTH1-transfected cells were treated with or without TCA (100 µM) for 24 h, and then cell viability, intracellular iron level, MDA level, and 4-HNE level were detected, mean ± SD, n = 5; *p &lt; 0.05 vs. Control, **p &lt; 0.01 vs. Control, #p &lt; 0.05 vs. TCA, ##p &lt; 0.01vs. TCA, †p &lt; 0.05 OE + TCA vs. NC + TCA, ††p &lt; 0.01 OE + TCA vs. NC + TCA. D. Untransfected cells and FTH1-transfected cells were treated with or without TCA (100 µM) for 24 h, and then ROS level was detected by flow cytometry E. Representative images of HTR-8/SVneo cells stained with DCFH-DA and MitoSox-Red. Scale bar, 5 μm. FTH1: Ferritin Heavy Chain 1, OE: FTH1 overexpression, NC: negative control, KRAS: GTPase KRas, GPX4: glutathione peroxidase 4, SLC7A11: solute carrier family 7 member 11, TCA: taurocholic acid, DFO: deferoxamine, OD: optical density, MDA: malondialdehyde,4-HNE: lipid peroxidation, ROS: reactive oxygen species, DCFH-DA: 2′,7′-dichlorofluorescein diacetate</p> <p>For both the NC and OE groups, TCA could decrease the expression of FTH1, GPX4, SLC7A11, and KRAS (Fig. 5B). In contrast, co-treatment with DFO could inhibit these changes to some extent. The expression of KRAS was lower in the OE TCA group compared with that in the NC TCA group.</p> <p>As shown in Fig. 5C, the cell growth rate was higher in the OE group than in the NC group. Both the proliferation and viability of cells were remarkably inhibited in the TCA group compared with their corresponding control groups. However, cell proliferation increased after 24 h in TCA-induced cell co-treatment with the DFO group compared with that in the TCA group.</p> <p>The MDA, intracellular 4-HNE, and iron levels were significantly higher in the NC and OE groups treated with TCA (Fig. 5C) relative to their control groups, respectively. Contrarily, all indices were reduced in the TCA + DFO group compared with that in the NC and OE TCA groups. In the HTR-8/SVneo cells that overexpressed FTH1, the intracellular iron levels in the TCA group reduced up to 38% compared with that in the NC TCA group. In addition, the MDA levels in the OE TCA group decreased from 13.6 ± 0.15 nmol/mg to 9.93 ± 0.46 nmol/mg compared with that in the NC TCA group. The results of intracellular 4-HNE displayed a similar trend. The cROS and mROS levels were evaluated by confocal laser scanning with different fluorescence stains. ROS levels in the different groups were monitored using a DCFH-DA probe. As shown in Fig. 5D, the cells exhibited considerably increased DCFH-DA fluorescent intensity when treated with TCA. In contrast, DFO suppressed the trend. The fluorescent intensity in the TCA-induced OE group was reduced by nearly 50% compared with that in the TCA-induced NC group. As indicated in Fig. 5E, massive green fluorescence and red fluorescence distribution in the TCA-treated NC group and TCA-treated OE group were observed. These images were consistent with the results of ROS detection obtained via flow cytometry. The lower green fluorescence and red fluorescence distribution in the OE TCA group relative to the NC TCA group indicated the effective reduction of cROS and mROS levels, which represented the protective effect of overexpressed FTH1 on TCA-induced cell ferroptosis (Fig. 5E).</p> <hd id="AN0182154229-23">Discussion</hd> <p>The placenta is a critical organ for the maintenance of fetal development and growth in utero and is the only medium for the exchange of nutrients and oxygen between mother and fetus during pregnancy [[<reflink idref="bib27" id="ref40">27</reflink>]]. Thus, the placental tissues from patients with ICP were used to verify the expression of FTH1. In our study, the mRNA and protein levels revealed significant downregulation of FTH1 (Fig. 1). Based on the results of the in vitro model of ICP, TCA promoted HTR-8/SVneo and EAhy926 cell death in a dose-dependent manner (Fig. 2A), ultimately aligning with the results of previous studies [[<reflink idref="bib4" id="ref41">4</reflink>], [<reflink idref="bib24" id="ref42">24</reflink>], [<reflink idref="bib26" id="ref43">26</reflink>], [<reflink idref="bib28" id="ref44">28</reflink>]]. As for the cell viability of HTR-8/SVneo cells treated with TCA(100 µM) for 48 h increased compared with that of treated with TCA(50 µM), the propensities of cells exposed to a high concentration of TCA for more than 48 h may lead to specific resistance mechanisms. Further, the degradation of TCA during long-term treatment could serve as another reason. Thus shortened treatment hours may be more reasonable. Collectively, HTR-8/SVneo cells treated with TCA (100 µM) for 24 h were selected to represent the pathological environment of ICP in our study. FTH1 was also found to be significantly downregulated in the TCA-induced HTR-8/SVneo cells (Fig. 2C). Notably, a high concentration of TCA in serum could cause placental damage and the etiology of fetal complications in patients with ICP. Trophoblast dysfunction was caused by TCA, leading to the inhibition of physiologic transfer of fetal bile acids to the mother [[<reflink idref="bib5" id="ref45">5</reflink>], [<reflink idref="bib10" id="ref46">10</reflink>]]. Consequently, the accumulation of fetal bile acids in fetus could explain the adverse pregnancy outcomes of ICP.</p> <p>Apoptosis and the cell cycle were previously evaluated in TCA-induced HTR-8/SVneo cells and could be involved in the development of ICP [[<reflink idref="bib28" id="ref47">28</reflink>]]; however, ferroptosis in ICP has not been examined. GPX4 as a unique intracellular antioxidant enzyme it is, could decrease lipid peroxidation of polyunsaturated fatty acids (PUFA) in cellular membranes and ferroptosis, it has been well documented that Gpx4 plays a vital role in the signal transduction of ferroptosis [[<reflink idref="bib30" id="ref48">30</reflink>]]. SLC7A11, the catalytic subunit of the cystine/glutamate antiporter system xc−, is the major transporter of extracellular cystine, also is a key role in maintaining intracellular glutathione levels and protecting cells from oxidative-stress-induced cell death, such as ferroptosis [[<reflink idref="bib13" id="ref49">13</reflink>]]. Thus GPX4 and SLC7A11 were considered as central regulators of ferroptosis, and the downregulation of GPX4 and SLC7A11 has always been regarded as markers of ferroptosis. As shown in Fig. 4A, TCA can cause ferroptosis. However, ferroptosis was largely offset by DFO.</p> <p>KRAS plays an important role in the regulation of cell proliferation and is a frequently mutated oncogene in cancers [[<reflink idref="bib32" id="ref50">32</reflink>]]. As shown in Fig. 3, KRAS could inhibit the expression of FTH1 through a transcriptional mechanism and the activation of organismal death. Thus, the expression of KRAS and the relationship with FTH1 were examined in our studies. The related mechanisms of FTH1 response to TCA treatment and the impact of FTH1 downregulation on ferroptosis induced by TCA in human trophoblast HTR-8/SVeno cells were explored. According to the study by Kiessling et al., the inhibition of constitutively active human NF-κB complex decreased the expression of FTH1 mRNA in T lymphocytes from patients with Sézary syndrome [[<reflink idref="bib33" id="ref51">33</reflink>]]. The mRNA of FTH1 was also downregulated by KRAS antisense in human pancreatic cancer cell lines AsPC-1 cells [[<reflink idref="bib34" id="ref52">34</reflink>]]. Based on the above studies, the expression of KRAS decreased in the TCA group (Fig. 4A) but remained suppressed in the TCA + DFO group. Such finding indicates that KRAS is not involved in ferroptosis, but TCA could suppress the expression of KRAS. Considering ferroptosis as a novel cell death form characterized by iron-dependent accumulation of lipid peroxidation [[<reflink idref="bib13" id="ref53">13</reflink>], [<reflink idref="bib19" id="ref54">19</reflink>], [<reflink idref="bib30" id="ref55">30</reflink>]], the intracellular iron, MDA, 4-HNE, and ROS levels were significantly increased in the TCA group (Fig. 4). As an end-product of lipid oxidation, MDA could cause metabolic disorders and indirectly reflect the degree of peroxidation [[<reflink idref="bib35" id="ref56">35</reflink>]]. Increased ROS and MDA levels can induce DNA damage, which could promote the development of ferroptosis. Cysteine deficiency-induced ferroptosis leads to hyperpolarization of the mitochondrial membrane and lipid peroxide accumulation [[<reflink idref="bib36" id="ref57">36</reflink>]]. In the TCA group, higher mROS levels were observed, which is consistent with the ROS levels (Fig. 4F, G). Each FTH1 can accommodate 4,500 ferrous iron atoms, which are then oxidized to ferric iron by FTH1 in an oxygen-dependent manner [[<reflink idref="bib19" id="ref58">19</reflink>], [<reflink idref="bib37" id="ref59">37</reflink>]]. The suppressed FTH1 in the TCA group can cause excessive iron release; thus, labile iron generates more ROS through the Fenton reaction or oxidation [[<reflink idref="bib35" id="ref60">35</reflink>], [<reflink idref="bib38" id="ref61">38</reflink>]]. On the other hand, lipoxygenase (LOX), which is the most important enzyme in PUFA metabolism, promotes the oxidation of PUFA [[<reflink idref="bib40" id="ref62">40</reflink>]]. Iron overload aggravates PUFA peroxidation as a catalysis of the LOXs. Free radicals attack membrane lipids, initiate the chain reaction of lipid peroxidation, and lead to cell damage. Eventually, a high concentration of TCA induces ferroptosis.</p> <p>Based on the findings obtained with the overexpressed FTH1 cells, FTH1 plays a key role in TCA-induced ferroptosis. In overexpressed FTH1 cells, the intracellular iron, MDA, 4-HNE, and ROS levels were significantly reduced compared with those in the NC group (Fig. 5C‒E). The overexpression of FTH1 in cells displayed a similar trend in response to DFO treatment. TCA-induced ferroptosis was inhibited by the overexpression of FTH1. The overexpressed FTH1 can lead to the storage of more ferrous iron atoms and their maintenance in a non-toxic form. FTH1 plays a vital role in the maintenance of the intracellular iron balance and deprivation of the ferrous iron atoms required for the Fenton reaction. Overexpressed FTH1 prevented damage caused by ROS, MDA, and 4-HNE in cells (Fig. 5). Collectively, the downregulation of FTH1 in patients with ICP could be the potential target for ICP through ferroptosis. Mechanistically, FTH1 is a critical mediator of TCA-induced ferroptosis in HTR-8/SVneo cells. Accordingly, the overexpression of FTH1 could prevent HTR-8/SVneo cell death owing to TCA-induced ferroptosis.</p> <p>Ursodeoxycholic acid (UDCA), cholestyramine, obeticholic acid (OCA), S-adenosyl methionine, dexamethasone, phenobarbital, and antihistamines, are used to treat ICP [[<reflink idref="bib41" id="ref63">41</reflink>]]. Recently, resveratrol, magnesium sulphate, and coenzyme Q10 were reported to be effective at relieving the symptoms of ICP [[<reflink idref="bib11" id="ref64">11</reflink>], [<reflink idref="bib26" id="ref65">26</reflink>], [<reflink idref="bib43" id="ref66">43</reflink>]]. Our findings support this notion as the compounds have antioxidant properties, such as reducing the intracellular levels of ROS or 4-HNE, and inhibiting the production of MAD to inhibit the development of TCA-induced ferroptosis in ICP. The current study highlighted the potential role of FTH1 in ICP. To our knowledge, this is the first report of TCA-induced ferroptosis in HTR-8/SVneo cells involving FTH1. However, the relationship between FTH1 and KRAS and how TCA affects FTH1 and KRAS ought to be determined in vitro and in vivo.</p> <hd id="AN0182154229-24">Conclusion</hd> <p>Pregnant women with ICP commonly have a high serum bile acid level. As the most predominant component of bile acid, TCA could suppress the expression of FTH1 in human trophoblast cells. The downregulation of FTH1 leads to a labile iron pool and excessive iron in cells, which contributes to the Fenton reaction. Consequently, ROS generated via the Fenton reaction and lipid peroxidation accumulation triggered ferroptosis. In summary, TCA induced cell death by triggering FTH1-dependent ferroptosis, which highlights a potential ferroptosis inducer for ICP treatment.</p> <hd id="AN0182154229-25">Acknowledgements</hd> <p>We would like to thank Xinrun Zhuang for his valuable discussion and assistance with the experiments.</p> <hd id="AN0182154229-26">Author contributions</hd> <p>WJ. Z. contributed to experiments and the draft of the manuscript. HJ. Y. contributed significantly to experiments and manuscript preparation. YJ. G. and MN. Y. helped perform the analysis with constructive discussions. MR. S. and SK. C. contributed to sample collection. W. G. and YY. H. contributed to the conception of the study and supervised the manuscript. All authors read and approved the final manuscript.</p> <hd id="AN0182154229-27">Funding</hd> <p>This study was funded by the National Key Research and Development Program of China (Grant Number: 2019YFA0802604) and National Natural Science Foundation of China (Grant Number: 81861128021). The funders had no role in the study design, data collection, data analysis, data interpretation, or writing of the report. The corresponding author had full access to all data in the study and made the final decision to submit the study for publication.</p> <hd id="AN0182154229-28">Data availability</hd> <p>Data and material used will be made available on request to corresponding author.</p> <hd id="AN0182154229-29">Declarations</hd> <p></p> <hd id="AN0182154229-30">Ethics approval and consent to participate</hd> <p>This study was performed according to the ethical guideline of the Institutional Ethics Committee of the International Peace Maternity and Child Health Hospital(IPMCH; approval number: [GKLW] 2019-14).The study was conducted in accordance with the Declaration of Helsinki.All participants provided written informed consents and the ethics committee approved the consent procedure.</p> <hd id="AN0182154229-31">Consent for publication</hd> <p>Not applicable.</p> <hd id="AN0182154229-32">Competing interests</hd> <p>The authors declare no competing interests.</p> <hd id="AN0182154229-33">Abbreviations</hd> <p></p> <p>• ICP</p> <p></p> <ulist> <item> Intrahepatic cholestasis of pregnancy</item> <p></p> </ulist> <p>• FTH1</p> <p></p> <ulist> <item> Ferritin Heavy Chain 1</item> <p></p> </ulist> <p>• TCA</p> <p></p> <ulist> <item> Taurocholic acid</item> <p></p> </ulist> <p>• TBA</p> <p></p> <ulist> <item> Total bile acid</item> <p></p> </ulist> <p>• IPMCH</p> <p></p> <ulist> <item> The International Peace Maternity and Child Health Hospital</item> <p></p> </ulist> <p>• IPA</p> <p></p> <ulist> <item> Ingenuity Pathway Analysis</item> <p></p> </ulist> <p>• CCK8</p> <p></p> <ulist> <item> Cell Counting Kit-8</item> <p></p> </ulist> <p>• MDA</p> <p></p> <ulist> <item> Malondialdehyde</item> <p></p> </ulist> <p>• 4-HNE</p> <p></p> <ulist> <item> Lipid peroxidation</item> <p></p> </ulist> <p>• ROS</p> <p></p> <ulist> <item> Reactive oxygen species</item> <p></p> </ulist> <p>• cROS</p> <p></p> <ulist> <item> Cellular ROS</item> <p></p> </ulist> <p>• DCFH-DA</p> <p></p> <ulist> <item> 2′,7′-dichlorofluorescein diacetate</item> <p></p> </ulist> <p>• mROS</p> <p></p> <ulist> <item> Mitochondrial ROS</item> <p></p> </ulist> <p>• OE</p> <p></p> <ulist> <item> FTH1 overexpression</item> <p></p> </ulist> <p>• NC</p> <p></p> <ulist> <item> Negative control</item> <p></p> </ulist> <p>• DFO</p> <p></p> <ulist> <item> Deferoxamine</item> <p></p> </ulist> <p>• OD</p> <p></p> <ulist> <item> Optical density</item> <p></p> </ulist> <p>• KRAS</p> <p></p> <ulist> <item> GTPase KRas</item> <p></p> </ulist> <p>• GPX4</p> <p></p> <ulist> <item> Glutathione peroxidase 4</item> <p></p> </ulist> <p>• SLC7A11</p> <p></p> <ulist> <item> Solute carrier family 7 member 11</item> <p></p> </ulist> <p>• PUFA</p> <p></p> <ulist> <item> Peroxidation of poly-unsaturated fatty acids</item> <p></p> </ulist> <p>• LOX</p> <p></p> <ulist> <item> Lipoxygenase</item> <p></p> </ulist> <p>• UDCA</p> <p></p> <ulist> <item> Ursodeoxycholic acid</item> <p></p> </ulist> <p>• OCA</p> <p></p> <ulist> <item> Obeticholic acid</item> </ulist> <hd id="AN0182154229-34">Electronic supplementary material</hd> <p>Below is the link to the electronic supplementary material.</p> <p>Graph: Supplementary Material 1</p> <p>Graph: Supplementary Material 2</p> <p>Graph: Supplementary Material 3</p> <p>Graph: Supplementary Material 4</p> <p>Graph: Supplementary Material 5</p> <p>Graph: Supplementary Material 6</p> <p>Graph: Supplementary Material 7</p> <hd id="AN0182154229-35">Publisher's note</hd> <p>Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.</p> <ref id="AN0182154229-36"> <title> References </title> <blist> <bibl id="bib1" idref="ref1" type="bt">1</bibl> <bibtext> Hague W, Bill, Williamson C, Beuers U. Intrahepatic cholestasis of pregnancy: introduction and overview 2024. Obstet Med. 2024; 17: 138-43. 10.1177/1753495X241265772. 39262909</bibtext> </blist> <blist> <bibl id="bib2" idref="ref5" type="bt">2</bibl> <bibtext> Roediger R, Fleckenstein J. Intrahepatic cholestasis of pregnancy. Clin Liver Dis 2024;23.</bibtext> </blist> <blist> <bibl id="bib3" idref="ref2" type="bt">3</bibl> <bibtext> Ahmed KT, Almashhrawi AA, Rahman RN, Hammoud GM, Ibdah JA. Liver diseases in pregnancy: diseases unique to pregnancy. World J Gastroenterol Nov. 2013; 21; 43: 7639-46. 10.3748/wjg.v19.i43.7639</bibtext> </blist> <blist> <bibl id="bib4" idref="ref6" type="bt">4</bibl> <bibtext> Williamson C, Geenes V. Intrahepatic cholestasis of pregnancy. Obstet Gynecol Jul. 2014; 124; 1: 120-33. 10.1097/aog.0000000000000346</bibtext> </blist> <blist> <bibl id="bib5" idref="ref3" type="bt">5</bibl> <bibtext> Gabzdyl EM, Schlaeger JM. Intrahepatic cholestasis of pregnancy: a critical clinical review. J Perinat Neonatal Nurs Jan-Mar. 2015; 29; 1: 41-50. 10.1097/jpn.0000000000000077</bibtext> </blist> <blist> <bibl id="bib6" idref="ref4" type="bt">6</bibl> <bibtext> Lee NM, Brady CW. Liver disease in pregnancy. World J Gastroenterol Feb. 2009; 28; 8: 897-906. 10.3748/wjg.15.897</bibtext> </blist> <blist> <bibl id="bib7" idref="ref13" type="bt">7</bibl> <bibtext> Diken Z, Usta IM, Nassar AH. A clinical approach to intrahepatic cholestasis of pregnancy. Am J Perinatol Jan. 2014; 31; 1: 1-8. 10.1055/s-0033-1333673</bibtext> </blist> <blist> <bibl id="bib8" idref="ref15" type="bt">8</bibl> <bibtext> Zhou Q, Yuan Y, Wang Y, He Z, Liang Y, Qiu S. The severity of intrahepatic cholestasis during pregnancy increases risks of adverse outcomes beyond stillbirth: evidence from 15,826 patients. BMC Pregnancy Childbirth. 2024; 24: 476. 10.1186/s12884-024-06645-2. 38997626. 11241884</bibtext> </blist> <blist> <bibl id="bib9" type="bt">9</bibl> <bibtext> Li X, Kong Y, Ren Y, Li Y, Xu J, Zhan Y. The impact of maternal intrahepatic cholestasis during pregnancy on the growth trajectory of offspring: a population-based nested case–control cohort study. BMC Pregnancy Childbirth. 2024; 24: 413. 10.1186/s12884-024-06559-z. 38849722. 11157880</bibtext> </blist> <blist> <bibtext> Wu WB, Menon R, Xu YY. Downregulation of peroxiredoxin-3 by hydrophobic bile acid induces mitochondrial dysfunction and cellular senescence in human trophoblasts. Sci Rep Dec 13. 2016; 6: 38946. 1:CAS:528:DC%2BC28XitFGjtrzJ. 10.1038/srep38946</bibtext> </blist> <blist> <bibtext> Martinefski MR, Rodriguez MR, Buontempo F, Lucangioli SE, Bianciotti LG, Tripodi VP. Coenzyme Q 10 supplementation: a potential therapeutic option for the treatment of intrahepatic cholestasis of pregnancy. Eur J Pharmacol Sep. 2020; 5: 173270. 1:CAS:528:DC%2BB3cXhtlSiu7jN. 10.1016/j.ejphar.2020.173270</bibtext> </blist> <blist> <bibtext> Dixon SJ, Lemberg KM, Lamprecht MR, Skouta R, Zaitsev EM, Gleason CE. Ferroptosis: an iron-dependent form of nonapoptotic cell death. Cell. 2012; 149; 5: 1060-72. 1:CAS:528:DC%2BC38XnslSntrw%3D. 10.1016/j.cell.2012.03.042. 22632970. 3367386</bibtext> </blist> <blist> <bibtext> Stockwell BR, Friedmann Angeli JP, Bayir H. Ferroptosis: a regulated cell death Nexus linking metabolism, Redox Biology, and Disease. Cell Oct. 2017; 5; 2: 273-85. 1:CAS:528:DC%2BC2sXhs1Wqs7%2FL. 10.1016/j.cell.2017.09.021</bibtext> </blist> <blist> <bibtext> Liang D, Feng Y, Zandkarimi F, Wang H, Zhang Z, Kim J. Ferroptosis surveillance independent of GPX4 and differentially regulated by sex hormones. Cell. 2023; 186: 2748-e276422. 1:CAS:528:DC%2BB3sXhtFCnsbvM. 10.1016/j.cell.2023.05.003. 37267948. 10330611</bibtext> </blist> <blist> <bibtext> Yang N, Wang Q, Ding B. Expression profles and functions of ferroptosis-related genes in the placental tissue samples of early- and late-onset preeclampsia patients.Yang et al. BMC Pregnancy Childbirth. 2022; 22: 87. 1:CAS:528:DC%2BB38XnsFyhsr4%3D. 10.1186/s12884-022-04423-6. 35100981. 8805258</bibtext> </blist> <blist> <bibtext> Muckenthaler MU, Rivella S, Hentze MW, Galy B. A Red Carpet for Iron Metabolism. Cell Jan. 2017; 26; 3: 344-61. 1:CAS:528:DC%2BC2sXhslaqsL4%3D. 10.1016/j.cell.2016.12.034</bibtext> </blist> <blist> <bibtext> Anderson SA, Nizzi CP, Chang YI. The IRP1-HIF-2α axis coordinates iron and oxygen sensing with erythropoiesis and iron absorption. Cell Metab Feb. 2013; 5; 2: 282-90. 1:CAS:528:DC%2BC3sXitFeqtbg%3D. 10.1016/j.cmet.2013.01.007</bibtext> </blist> <blist> <bibtext> Chen X, Yu C, Kang R, Tang D. Iron Metabolism in Ferroptosis. Front cell Dev Biology. 2020; 8: 590226. 10.3389/fcell.2020.590226</bibtext> </blist> <blist> <bibtext> Kong N, Chen X, Feng J. Baicalin induces ferroptosis in bladder cancer cells by downregulating FTH1. Acta Pharm Sinica B Dec. 2021; 11; 12: 4045-54. 1:CAS:528:DC%2BB3MXitFKis73K. 10.1016/j.apsb.2021.03.036</bibtext> </blist> <blist> <bibtext> Zhang Q, Chen C, Zou X, Wu W, Di Y, Li N. Iron promotes ovarian cancer malignancy and advances platinum resistance by enhancing DNA repair via FTH1/FTL/POLQ/RAD51 axis. Cell Death Dis. 2024; 15: 329. 1:CAS:528:DC%2BB2cXhtFehtb7I. 10.1038/s41419-024-06688-5. 38740757. 11091064</bibtext> </blist> <blist> <bibtext> Cui J, Chen Y, Yang Q, Zhao P, Yang M, Wang X et al. Protosappanin a protects DOX-Induced Myocardial Injury and Cardiac Dysfunction by Targeting ACSL4/FTH1 Axis‐Dependent Ferroptosis. Adv Sci 2024;:2310227.</bibtext> </blist> <blist> <bibtext> Yang Y, Wu Q, Shan X, Zhou H, Wang J, Hu Y. Ginkgolide B attenuates cerebral ischemia-reperfusion injury via inhibition of ferroptosis through disrupting NCOA4-FTH1 interaction. J Ethnopharmacol. 2024; 318: 116982. 1:CAS:528:DC%2BB3sXhslShu7fO. 10.1016/j.jep.2023.116982. 37532074</bibtext> </blist> <blist> <bibtext> Zeng W, Hou Y, Gu W. Proteomic biomarkers of Intrahepatic Cholestasis of pregnancy. Reprod Sci. 2024. 10.1007/s43032-023-01437-z. 39567465</bibtext> </blist> <blist> <bibtext> Qin X, Ni X, Mao X, Ying H, Du Q. Cholestatic pregnancy is associated with reduced VCAM1 expression in vascular endothelial cell of placenta. Reprod Toxicol Dec. 2017; 74: 23-31. 1:CAS:528:DC%2BC2sXhsVOhsLzI. 10.1016/j.reprotox.2017.08.002</bibtext> </blist> <blist> <bibtext> Geenes VL, Lim YH, Bowman N. A placental phenotype for intrahepatic cholestasis of pregnancy. Placenta. 2011; 32; 12: 1026-32. 1:CAS:528:DC%2BC3MXhsFGjt7jF. 10.1016/j.placenta.2011.09.006. 22015023</bibtext> </blist> <blist> <bibtext> Liao E, Li Z, Shao Y. Resveratrol regulates the silent information regulator 1-nuclear factor-κB signaling pathway in intrahepatic cholestasis of pregnancy. Hepatol Res Nov. 2018; 48; 12: 1031-44. 1:CAS:528:DC%2BC1cXitlSrtLrE. 10.1111/hepr.13198</bibtext> </blist> <blist> <bibtext> Maltepe E, Fisher SJ. Placenta: the forgotten organ. Annu Rev Cell Dev Biol. 2015; 31: 523-52. 1:CAS:528:DC%2BC2MXhs1WqtbvF. 10.1146/annurev-cellbio-100814-125620. 26443191</bibtext> </blist> <blist> <bibtext> Zhang T, Zhao C, Luo L. High concentraction of taurocholic acid induced apoptosis in HTR-8/SVneo cells via overexpression of ERp29 and activation of p38. Placenta Jul. 2014; 35; 7: 496-500. 1:CAS:528:DC%2BC2cXnsVajsL8%3D. 10.1016/j.placenta.2014.03.023</bibtext> </blist> <blist> <bibtext> Zou S, Zou P, Wang Y. ERp29 inhibition attenuates TCA toxicity via affecting p38/p53- dependent pathway in human trophoblast HTR-8/SVeno cells. Arch Biochem Biophys Nov. 2019; 15: 108125. 1:CAS:528:DC%2BC1MXhvFert73K. 10.1016/j.abb.2019.108125</bibtext> </blist> <blist> <bibtext> Doll S, Proneth B, Tyurina YY. ACSL4 dictates ferroptosis sensitivity by shaping cellular lipid composition. Nat Chem Biol Jan. 2017; 13; 1: 91-8. 1:CAS:528:DC%2BC28XhvVGgtLrL. 10.1038/nchembio.2239</bibtext> </blist> <blist> <bibtext> Kagan VE, Mao G, Qu F. Oxidized arachidonic and adrenic PEs navigate cells to ferroptosis. Nat Chem Biol Jan. 2017; 13; 1: 81-90. 1:CAS:528:DC%2BC28XhvVGgtLrE. 10.1038/nchembio.2238</bibtext> </blist> <blist> <bibtext> Pandey D, Chauhan SC, Kashyap VK, Roy KK. Structural insights into small-molecule KRAS inhibitors for targeting KRAS mutant cancers. Eur J Med Chem. 2024; 277: 116771. 1:CAS:528:DC%2BB2cXhvVWisrrM. 10.1016/j.ejmech.2024.116771. 39167893</bibtext> </blist> <blist> <bibtext> Kiessling MK, Klemke CD, Kaminski MM, Galani IE, Krammer PH, Gülow K. Inhibition of constitutively activated nuclear factor-kappab induces reactive oxygen species- and iron-dependent cell death in cutaneous T-cell lymphoma. Cancer Res Mar. 2009; 15; 6: 2365-74. 10.1158/0008-5472.Can-08-3221</bibtext> </blist> <blist> <bibtext> Ohnami S, Matsumoto N, Nakano M. Identification of genes showing differential expression in antisense K-ras-transduced pancreatic cancer cells with suppressed tumorigenicity. Cancer Res Nov. 1999; 1; 21: 5565-71</bibtext> </blist> <blist> <bibtext> Yang M, Chen P, Liu J. Clockophagy is a novel selective autophagy process favoring ferroptosis. Sci Adv Jul. 2019; 5; 7: eaaw2238. 1:CAS:528:DC%2BB3cXhtlakur%2FK. 10.1126/sciadv.aaw2238</bibtext> </blist> <blist> <bibtext> Gao M, Yi J, Zhu J. Role of Mitochondria in Ferroptosis. Mol Cell Jan. 2019; 17; 2: 354-e3633. 1:CAS:528:DC%2BC1cXisF2ks7fI. 10.1016/j.molcel.2018.10.042</bibtext> </blist> <blist> <bibtext> Carmona U, Li L, Zhang L, Knez M. Ferritin light-chain subunits: key elements for the electron transfer across the protein cage. Chem Commun (Camb) Dec. 2014; 18; 97: 15358-61. 10.1039/c4cc07996e</bibtext> </blist> <blist> <bibtext> Teschke R, Aluminum. Arsenic, Beryllium, Cadmium, Chromium, Cobalt, copper, Iron, lead, Mercury, Molybdenum, Nickel, Platinum, Thallium, Titanium, Vanadium, and zinc: Molecular aspects in Experimental Liver Injury. IJMS. 2022; 23: 12213. 1:CAS:528:DC%2BB38Xisl2ntLfF. 10.3390/ijms232012213. 36293069. 9602583</bibtext> </blist> <blist> <bibtext> Sies H, Belousov VV, Chandel NS, Davies MJ, Jones DP, Mann GE. Defining roles of specific reactive oxygen species (ROS) in cell biology and physiology. Nat Rev Mol Cell Biol. 2022; 23: 499-515. 1:CAS:528:DC%2BB38XksFCnsLw%3D. 10.1038/s41580-022-00456-z. 35190722</bibtext> </blist> <blist> <bibtext> Navratil AR, Shchepinov MS, Dennis EA. Lipidomics reveals dramatic physiological Kinetic Isotope effects during the enzymatic oxygenation of Polyunsaturated fatty acids Ex vivo. J Am Chem Soc Jan. 2018; 10; 1: 235-43. 1:CAS:528:DC%2BC2sXhvFaqs7jJ. 10.1021/jacs.7b09493</bibtext> </blist> <blist> <bibtext> Ozkan S, Ceylan Y, Ozkan OV, Yildirim S. Review of a challenging clinical issue: Intrahepatic cholestasis of pregnancy. World J Gastroenterol Jun. 2015; 21; 23: 7134-41. 10.3748/wjg.v21.i23.7134</bibtext> </blist> <blist> <bibtext> Wu JS, Liu Q, Fang SH. Quantitative proteomics reveals the Protective effects of Huangqi Decoction against Acute Cholestatic Liver Injury by inhibiting the NF-κB/IL-6/STAT3 signaling pathway. J Proteome Res Feb. 2020; 7; 2: 677-87. 1:CAS:528:DC%2BC1MXisVekt7rK. 10.1021/acs.jproteome.9b00563</bibtext> </blist> <blist> <bibtext> Han F, Xu L, Huang Y, Chen T, Zhou T, Yang L. Magnesium sulphate can alleviate oxidative stress and reduce inflammatory cytokines in rat placenta of intrahepatic cholestasis of pregnancy model. Arch Gynecol Obstet Sep. 2018; 298; 3: 631-8. 1:CAS:528:DC%2BC1cXhtlKmu7zO. 10.1007/s00404-018-4850-1</bibtext> </blist> </ref> <aug> <p>By Wei-jian Zeng; Hua-jing Yang; Ying-jie Gu; Meng-nan Yang; Meng-ru Sun; Sheng-kai Cheng; Yan-yan Hou and Wei Gu</p> <p>Reported by Author; Author; Author; Author; Author; Author; Author; Author</p> </aug> <nolink nlid="nl1" bibid="bib10" firstref="ref16"></nolink> <nolink nlid="nl2" bibid="bib11" firstref="ref17"></nolink> <nolink nlid="nl3" bibid="bib12" firstref="ref18"></nolink> <nolink nlid="nl4" bibid="bib13" firstref="ref19"></nolink> <nolink nlid="nl5" bibid="bib14" firstref="ref20"></nolink> <nolink nlid="nl6" bibid="bib15" firstref="ref21"></nolink> <nolink nlid="nl7" bibid="bib16" firstref="ref22"></nolink> <nolink nlid="nl8" bibid="bib17" firstref="ref23"></nolink> <nolink nlid="nl9" bibid="bib18" firstref="ref24"></nolink> <nolink nlid="nl10" bibid="bib19" firstref="ref25"></nolink> <nolink nlid="nl11" bibid="bib21" firstref="ref26"></nolink> <nolink nlid="nl12" bibid="bib22" firstref="ref27"></nolink> <nolink nlid="nl13" bibid="bib23" firstref="ref28"></nolink> <nolink nlid="nl14" bibid="bib24" firstref="ref34"></nolink> <nolink nlid="nl15" bibid="bib25" firstref="ref37"></nolink> <nolink nlid="nl16" bibid="bib26" firstref="ref38"></nolink> <nolink nlid="nl17" bibid="bib27" firstref="ref40"></nolink> <nolink nlid="nl18" bibid="bib28" firstref="ref44"></nolink> <nolink nlid="nl19" bibid="bib30" firstref="ref48"></nolink> <nolink nlid="nl20" bibid="bib32" firstref="ref50"></nolink> <nolink nlid="nl21" bibid="bib33" firstref="ref51"></nolink> <nolink nlid="nl22" bibid="bib34" firstref="ref52"></nolink> <nolink nlid="nl23" bibid="bib35" firstref="ref56"></nolink> <nolink nlid="nl24" bibid="bib36" firstref="ref57"></nolink> <nolink nlid="nl25" bibid="bib37" firstref="ref59"></nolink> <nolink nlid="nl26" bibid="bib38" firstref="ref61"></nolink> <nolink nlid="nl27" bibid="bib40" firstref="ref62"></nolink> <nolink nlid="nl28" bibid="bib41" firstref="ref63"></nolink> <nolink nlid="nl29" bibid="bib43" firstref="ref66"></nolink> CustomLinks: – Url: https://resolver.ebsco.com/c/xy5jbn/result?sid=EBSCO:edsdoj&genre=article&issn=14712393&ISBN=&volume=25&issue=1&date=20250101&spage=1&pages=1-13&title=BMC Pregnancy and Childbirth&atitle=High%20taurocholic%20acid%20concentration%20induces%20ferroptosis%20by%20downregulating%20FTH1%20expression%20in%20intrahepatic%20cholestasis%20of%20pregnancy&aulast=Wei-jian%20Zeng&id=DOI:10.1186/s12884-025-07143-9 Name: Full Text Finder (for New FTF UI) (s8985755) Category: fullText Text: Find It @ SCU Libraries MouseOverText: Find It @ SCU Libraries – Url: https://doaj.org/article/f93066ee320141cf9cb53e89f1e12d49 Name: EDS - DOAJ (s8985755) Category: fullText Text: View record from DOAJ MouseOverText: View record from DOAJ |
|---|---|
| Header | DbId: edsdoj DbLabel: Directory of Open Access Journals An: edsdoj.f93066ee320141cf9cb53e89f1e12d49 RelevancyScore: 1064 AccessLevel: 3 PubType: Academic Journal PubTypeId: academicJournal PreciseRelevancyScore: 1064.22692871094 |
| IllustrationInfo | |
| Items | – Name: Title Label: Title Group: Ti Data: High taurocholic acid concentration induces ferroptosis by downregulating FTH1 expression in intrahepatic cholestasis of pregnancy – Name: Author Label: Authors Group: Au Data: <searchLink fieldCode="AR" term="%22Wei-jian+Zeng%22">Wei-jian Zeng</searchLink><br /><searchLink fieldCode="AR" term="%22Hua-jing+Yang%22">Hua-jing Yang</searchLink><br /><searchLink fieldCode="AR" term="%22Ying-jie+Gu%22">Ying-jie Gu</searchLink><br /><searchLink fieldCode="AR" term="%22Meng-nan+Yang%22">Meng-nan Yang</searchLink><br /><searchLink fieldCode="AR" term="%22Meng-ru+Sun%22">Meng-ru Sun</searchLink><br /><searchLink fieldCode="AR" term="%22Sheng-kai+Cheng%22">Sheng-kai Cheng</searchLink><br /><searchLink fieldCode="AR" term="%22Yan-yan+Hou%22">Yan-yan Hou</searchLink><br /><searchLink fieldCode="AR" term="%22Wei+Gu%22">Wei Gu</searchLink> – Name: TitleSource Label: Source Group: Src Data: BMC Pregnancy and Childbirth, Vol 25, Iss 1, Pp 1-13 (2025) – Name: Publisher Label: Publisher Information Group: PubInfo Data: BMC, 2025. – Name: DatePubCY Label: Publication Year Group: Date Data: 2025 – Name: Subset Label: Collection Group: HoldingsInfo Data: LCC:Gynecology and obstetrics – Name: Subject Label: Subject Terms Group: Su Data: <searchLink fieldCode="DE" term="%22Ferroptosis%22">Ferroptosis</searchLink><br /><searchLink fieldCode="DE" term="%22FTH1%22">FTH1</searchLink><br /><searchLink fieldCode="DE" term="%22Intrahepatic+cholestasis+of+pregnancy%22">Intrahepatic cholestasis of pregnancy</searchLink><br /><searchLink fieldCode="DE" term="%22Placenta%22">Placenta</searchLink><br /><searchLink fieldCode="DE" term="%22Taurocholic+acid%22">Taurocholic acid</searchLink><br /><searchLink fieldCode="DE" term="%22Gynecology+and+obstetrics%22">Gynecology and obstetrics</searchLink><br /><searchLink fieldCode="DE" term="%22RG1-991%22">RG1-991</searchLink> – Name: Abstract Label: Description Group: Ab Data: Abstract Background Intrahepatic cholestasis of pregnancy (ICP) is the most common liver disorder associated with pregnancy and is usually diagnosed based on high serum bile acid. However, the pathogenesis of ICP is unclear. Ferroptosis has been reported as an iron-dependent mechanism of cell death. Although the role of Ferritin Heavy Chain 1 (FTH1) in ferroptosis has been extensively studied in various diseases, its mechanism in ICP through ferroptosis is yet to be analyzed. Methods Placental tissues from patients with ICP and healthy controls were employed to verify the expression of FTH1. Taurocholic acid (TCA)-induced HTR-8/SVneo cells were established as an in vitro model for ICP, and ferroptosis-related experiments were performed. In particular, HTR-8/SVneo cells with or without overexpressing FTH1 and HTR-8/SVneo cells with or without TCA induction were investigated to explore the relationship between FTH1 and ferroptosis during ICP in vitro, respectively. Results FTH1 was significantly downregulated in the ICP group compared with the control group. Furthermore, FTH1 and ferroptosis-related protein levels were downregulated, while the intracellular iron, reactive oxygen species, and lipid peroxidation levels were upregulated in the TCA-induced HTR-8/SVneo cells. In contrast, ferroptosis was inhibited by overexpression of FTH1 in TCA-induced HTR-8/SVneo cells. Conclusions A high concentration of TCA in HTR-8/SVneo cells decreased the expression of FTH1. Overexpression of FTH1 could prevent cell death from ferroptosis induced by TCA. Thus, inhibiting the downregulation of FTH1 could be a potential therapeutic target for ICP treatment. – Name: TypeDocument Label: Document Type Group: TypDoc Data: article – Name: Format Label: File Description Group: SrcInfo Data: electronic resource – Name: Language Label: Language Group: Lang Data: English – Name: ISSN Label: ISSN Group: ISSN Data: 1471-2393 – Name: NoteTitleSource Label: Relation Group: SrcInfo Data: https://doaj.org/toc/1471-2393 – Name: DOI Label: DOI Group: ID Data: 10.1186/s12884-025-07143-9 – Name: URL Label: Access URL Group: URL Data: <link linkTarget="URL" linkTerm="https://doaj.org/article/f93066ee320141cf9cb53e89f1e12d49" linkWindow="_blank">https://doaj.org/article/f93066ee320141cf9cb53e89f1e12d49</link> – Name: AN Label: Accession Number Group: ID Data: edsdoj.f93066ee320141cf9cb53e89f1e12d49 |
| PLink | https://login.libproxy.scu.edu/login?url=https://search.ebscohost.com/login.aspx?direct=true&site=eds-live&scope=site&db=edsdoj&AN=edsdoj.f93066ee320141cf9cb53e89f1e12d49 |
| RecordInfo | BibRecord: BibEntity: Identifiers: – Type: doi Value: 10.1186/s12884-025-07143-9 Languages: – Text: English PhysicalDescription: Pagination: PageCount: 13 StartPage: 1 Subjects: – SubjectFull: Ferroptosis Type: general – SubjectFull: FTH1 Type: general – SubjectFull: Intrahepatic cholestasis of pregnancy Type: general – SubjectFull: Placenta Type: general – SubjectFull: Taurocholic acid Type: general – SubjectFull: Gynecology and obstetrics Type: general – SubjectFull: RG1-991 Type: general Titles: – TitleFull: High taurocholic acid concentration induces ferroptosis by downregulating FTH1 expression in intrahepatic cholestasis of pregnancy Type: main BibRelationships: HasContributorRelationships: – PersonEntity: Name: NameFull: Wei-jian Zeng – PersonEntity: Name: NameFull: Hua-jing Yang – PersonEntity: Name: NameFull: Ying-jie Gu – PersonEntity: Name: NameFull: Meng-nan Yang – PersonEntity: Name: NameFull: Meng-ru Sun – PersonEntity: Name: NameFull: Sheng-kai Cheng – PersonEntity: Name: NameFull: Yan-yan Hou – PersonEntity: Name: NameFull: Wei Gu IsPartOfRelationships: – BibEntity: Dates: – D: 01 M: 01 Type: published Y: 2025 Identifiers: – Type: issn-print Value: 14712393 Numbering: – Type: volume Value: 25 – Type: issue Value: 1 Titles: – TitleFull: BMC Pregnancy and Childbirth Type: main |
| ResultId | 1 |